The present invention relates to novel polynucleotides and proteins encoded by such polynucleotides, along with therapeutic, diagnostic, and research utilities for these polynucleotides and proteins. In particular, the invention relates to polynucleotides and proteins encoded by such polynucleotides that bind to xcex2-amyloid peptide, one of the primary components of amyloid deposits associated with Alzheimer""s disease.
Alzheimer""s disease (AD) is a progressive dementing disorder of the elderly characterized by a series of structural abnormalities of the brain. Neurons in multiple regions of the central nervous system (CNS) become dysfunctional and die, resulting in alterations in synaptic inputs. Cell bodies and proximal dendrites of these vulnerable neurons contain neurofibrillary angles composed of paired helical filaments, the main component of which is a phosphorylated microtubular-binding protein, namely tau. One of the hallmarks of the disease is the accumulation of amyloid containing deposits within the brain called senile (or neuritic) plaques. The principal component of amyloid plaques is xcex2-amyloid peptide (hereinafter xe2x80x9cBAP,xe2x80x9d also referred in the literature as Axcex2, xcex2AP, etc.), which forms dense aggregates during the course of AD.
BAP is a 39-43 amino acid peptide derived by proteolytic cleavage of amyloid precursor protein (hereinafter xe2x80x9cAPPxe2x80x9d) and composed of a portion of the transmembrane domain and the lumina/extracellular domain of APP. It is thought that the BAP peptide comprising 42 amino acids (BAP42) is potentially the more toxic aggregated form in humans. APP occurs as several BAP-containing isoforms. The major forms are comprised of 695, 751, and 770 amino acids, with the latter two APP containing a domain that shares structural and functional homologies with Kunitz serine protease inhibitors. In normal individuals, BAP does not accumulate and is rapidly removed from circulating fluids. However, the peptide can form plaques on surfaces of dystrophic dentrites and axons, microglia, and reactive astrocytes. The aggregation and deposition of BAP in neuritic plaques is postulated as one of the initiating events of AD. Investigation of the events leading to the expression and consequences of BAP and their individual roles in AD is a major focus of neuroscience research. In particular, the discovery of proteins that bind to BAP is critical to advance understanding of the pathogenesis of the disease and to potentially introduce novel therapeutic targets.
Until the present invention, proteins and fragments thereof that bind with human BAP and that may be involved in the biological effects of BAP in AD had not been identified.
This invention provides novel isolated polynucleotides that encode gene products that selectively bind human xcex2-amyloid peptide (BAP) amino acid sequences.
In one embodiment, the present invention provides a composition comprising an isolated polynucleotide selected from the group consisting of:
(a) polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1;
(b) a polynucleotide comprising the nucleotide sequence of a xcex2-amyloid peptide-binding protein (BBP) of clone BBP1-fl deposited under accession number ATCC 98617;
(c) a polynucleotide encoding a xcex2-amyloid peptide-binding protein (BBP) encoded by the cDNA insert of clone BBP1-fl deposited under accession number ATCC 98617;
(d) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1 from nucleotide 202 to nucleotide 807;
(e) a polynucleotide comprising the nucleotide sequence of a xcex2-amyloid peptide-binding protein (BBP) of clone pEK196 deposited under accession number ATCC 98399;
(f) a polynucleotide encoding a xcex2-amyloid peptide-binding protein (BBP) encoded by the cDNA insert of clone pEK196 deposited under accession number ATCC 98399;
(g) a polynucleotide encoding a protein comprising the amino acid sequence of SEQ ID NO: 2;
(h) a polynucleotide encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 2 having human xcex2-amyloid peptide-binding activity, the fragment comprising the amino acid sequence from amino acid 68 to amino acid 269 of SEQ ID NO: 2;
(i) a polynucleotide which is an allelic variant of the polynucleotide of (a)-(f) above:
(j) a polynucleotide which encodes a species homologue of the protein of (g)-(i) above; and
(k) a polynucleotide capable of hybridizing under stringent conditions to any one of the polynucleotides specified in (a)-(h).
Preferably such polynucleotide comprises the nucleotide sequence of SEQ ID NO: 1; the nucleotide sequence of a xcex2-amyloid peptide-binding protein (BBP) of clone BBP1-fl deposited under accession number ATCC 98617; or a polynucleotide encoding a xcex2-amyloid peptide-binding protein (BBP) encoded by the cDNA insert of clone BBP1-fl deposited under accession number ATCC 98617. Another embodiment provides the gene corresponding to the cDNA sequence of SEQ ID NO: 1.
In other embodiments, the present invention provides a composition comprising a protein, wherein said protein comprises an amino acid sequence selected from the group consisting of:
(a) the amino acid sequence of SEQ ID NO: 2;
(b) the amino acid sequence of SEQ ID NO: 2 from amino acid 68 to amino acid 269;
(c) the amino acid sequence encoded by the cDNA insert of clone BBP1-fl deposited under accession number ATCC 98617; and
(d) fragments of the amino acid sequence of SEQ ID NO: 2 comprising the amino acid sequence from amino acid 185 to amino acid 217 of SEQ ID NO: 2.
Preferably such protein comprises the amino acid sequence of SEQ ID NO: 2 or the amino acid sequence of SEQ ID NO: 2 from amino acid 68 to amino acid 269. Fusion proteins are also claimed in the present invention.
In certain preferred embodiments, the polynucleotide is operably lined to an expression control sequence. The invention also provides a host cell, including bacterial, yeast, insect, and mammalian cells, transformed with such polynucleotide compositions.
Processes are also provided for producing a BBP which comprises (a) growing a culture of the host cell of claim 3 in a suitable culture medium; and (b) purifying the protein from the culture medium.
Compositions comprising an antibody which specifically reacts with such BBPs are also provided by the present invention.
Methods and diagnostic processes are provided for detecting a disease state characterized by the aberrant expression of human BAP, as well as methods for identifying compounds that regulate the activity of BBPs.
Another embodiment of the invention includes transgenic animals comprising a polynucleotide encoding a BBP operably linked to an expression control sequence.
A further embodiment of the invention provides knockout animals in which the BBP1 gene has been functionally disrupted. The invention also relates to conditional knockout animals in which the BBP1 gene is disrupted in a temporal or tissue-specific manner or in which the BBP1 disruption can be induced by external stimuli.